Macromolecular MRI contrast agents based upon dendrimers obviate many deficiencies of serum albumin or linear polymer based MRI contrast agents. This is due to their preparation through an iterative synthesis methodology that promotes controlled size and shape of the dendrimer concomitantly generating the means for reproducible chemistry key to clinical translation. To create MRI contrast agents with dendrimers, the terminal primary amines of dendrimers are modified with chelated Gd(III) technology developed in the Chemistry Section. Historically, these reagents demonstrated a molar relaxivity 6 times that of Magnevist, the currently approved MRI contrast agent as reported in a seminal collaborative publication from the Chemistry Section. Macromolecular chelate conjugated dendrimer based Gd(III) MR contrast agents can be tuned for various applications by adjusting fundamental criteria: generation (MW & physical size), core elements (lipophilicity & charge), PEG conjugation (prolonging circulation minimizing non-specific organ uptake), lysine co-administration (assisting in renal clearance), and conjugation to targeting vectors (molecular targeting). Dendrimer based agents have also been selectively targeted, not only by conjugation to antibodies, but by other vectors, such as avidin to deliver exceptionally high levels of Gd(III) into disseminated intraperitoneal ovarian cancer tumor. This was done in conjunction with an optical imaging agent in parallel with our creation of multi-modality dendrimer based imaging agents. The incorporation of a NIR optical imaging dye into the MRI agent added an enhanced level of sensitivity to complement the resolution of the MRI imaging and provided an additional level of sensitivity for the imaging of lymphatics and sentinel nodes that can be envisioned as being translated to an intraoperative scenario wherein MRI imaging and mapping would supplement real-time surgical intervention and excision of malignancy. While the chemistry established the ability to create such macromolecular agents, the imaging resulted in compromised targeting which defined that these agents require very careful systematic investigation combined with equally careful defined characterization. Chelation chemistry for conjugation of Gd(III) complexes to dendrimer has moved from aqueous chemistry to organic solvents to enhance both characterization and consistency of yields. This chemistry has also evolved specialized analogs of established bifunctional chelation agents (patent filed) to address development of site-specific conjugation chemistry required for actively targeted dendrimer based imaging agents e.g, maleimides targeting a unique thiol residue, or agents functionalized with alkyne or azide groups for click chemistry conjugation strategies. NFS related Gd(III) toxicity resultant from use of less than adequately stable MRI contrast agents combined with low renal excretion prompted a new directionality in the choice of bifunctional chelating agent towards the use of DOTA as the bifunctional chelating agent. With the Gd(III) complex pre-formed prior to conjugation to all targeting vectors, toxicity concerns were traversed. This effort and result was adopted into all MR contrast projects exclusively using a pre-complexation of the Gd(III) conjugate in DOTA strategy also eliminating characterization complexity resulting from the creation of exceedingly difficult to characterize mixtures of products that limited reproducibility complicating clinical translation. Results validated this transformation as a successful strategy and also provided a far greater molar relaxivity; a 5-fold enhancement over the prior technology was reported while concurrently decreasing the actual physical amount of Gd(III) conjugated to the dendrimer by 65% further increasing the safety margin. The impact of this result extends across to all macromolecular MR contrast agents regardless of platform to fully address safety, characterization, and reproducibility thereby furthering an entire fields potential for clinical translation of such agents. The exquisite advantages of dendrimer based agents over low molecular weight agents, e.g. Magnevist, continues to be very clearly demonstrated. This improvement abrogating toxicity concerns that resulted in superior dendrimer based agents provided the chemistry to finally move forward with actively targeted dendrimer based contrast agents that are discrete characterized agents. This is being achieved through conjugation of a cystamine core dendrimer derived dendron conjugated in a 1:1 form with an antibody fragment. While successful, both isolation and purification continue to prove to be challenging with the dendrimer generation employed as well as relatively lower relaxivity results indicating the need to move onwards to a higher dendrimer generation. This investigation continues in parallel with the creation of a PEG appended trifunctional imaging agent that will permit incorporation of a radiological probe (PET imaging) with an optical probe. Recent studies related to the establishment of fully characterized and purified dye conjugates of monoclonal antibodies, e.g., indocyanine green conjugated to panitumumab targeting EGFR revealed that the prior literature was in fact faulty in its report of the identity of the agent as well as its mode of action calling into question later clinical utilization of that same agent. Our published work really calls into serious question the simplicity of optical imaging overriding the actual need for understanding what the agents being generated really are and their safe use. This has strongly demonstrated a real need for understanding vs. the ease of use driving these agents into clinical use.